Article Figures & Data
Figures
- FIGURE 1.
Maxadilan modulates TNF-α and IL-6 release by macrophages stimulated with LPS. BALB/c macrophages were preincubated for 2 h with different doses of maxadilan. An amount equal to 500ng/ml of LPS was then added to the macrophage monolayers and, 2 h (A) or 4 h (B) later, cell-free supernatants were collected. TNF-α and IL-6 contents were determined by ELISA. Values are shown as percent of LPS control from three independent experiments, expressed as mean ± SEM. * p < 0.05 compared with LPS alone. Levels of TNF-α in LPS control cultures varied from 100 pg/ml to 3 ng/ml, and of IL-6, from 1 to 6 ng/ml.
- FIGURE 2.
Maxadilan induces IL-6 release by normal macrophages. BALB/c macrophages were incubated for 6 h with the indicated doses of recombinant maxadilan. Supernatants were collected and tested for IL-6 by ELISA. Data represent the mean ± SEM of three separate experiments.
- FIGURE 3.
The inhibition of TNF-α by maxadilan on LPS-stimulated macrophages is blocked by indomethacin but not by anti IL-10. Macrophages were plated as described in Materials and Methods. After 2 h preincubation in the presence or in the absence of anti-IL-10 (5 μg/ml), indomethacin (1 μg/ml), and maxadilan (10 ng/ml), LPS (500 ng/ml) was added for an additional 4 h. Supernatants were collected and tested for TNF-α by ELISA. Results shown are from one representative of four independent experiments performed. Values are expressed as mean ± SEM of four determinations.
- FIGURE 4.
Maxadilan increases PGE2 from macrophages stimulated by LPS. BALB/c macrophages were preincubated for 2 h with different doses of maxadilan. An amount equal to 500 ng/ml of LPS was then added to the macrophage monolayers, and, 2 h later, the cell-free supernatants were collected and tested for the presence of PGE2 by enzyme immunoassay. Values are shown as percent of LPS control from four independent experiments, expressed as mean ± SEM. * p < 0.05 compared with LPS alone. Levels of PGE2 in LPS control cultures varied from 100 pg/ml to 1.5 ng/ml.
- FIGURE 5.
PACAP 6–38 blocks the effects of maxadilan on macrophages. BALB/c macrophages were preincubated for 2 h with maxadilan (10 ng/ml) in the presence or absence of PACAP 6–38 (1 μg/ml). LPS (500 ng/ml) was then added and the supernatants were collected after 2 or 4 h for TNF-α (A) and IL-6 (B), respectively, and tested by ELISA. Results shown are from one representative of five independent experiments performed. Values represent mean ± SD of four determinations.
- FIGURE 6.
Maxadilan dramatically enhances the level of intracellular cAMP in macrophages. A, Time course of cAMP accumulation. BALB/c macrophages were incubated with 10 ng/ml of maxadilan for different times, and the levels of intracellular cAMP were determined (techniques in Material and Methods). B, Dose response of cAMP accumulation. BALB/c macrophages were incubated with different doses of maxadilan for 15 min, and the levels of intracellular cAMP were determined. The figure is representative of three independent experiments.
Tables
Experiment 1 2 3 4 TNF-α (ng/ml) Medium 0 0 0 0 LPS 0.34 0.16 1.13 0.55 PACAP 0 0 0 0 PACAP + LPS 0.13 0.04 0.70 0.20 IL-6 (ng/ml) Medium 0 0 0 0 LPS 1.99 5.36 1.61 2.96 PACAP 0.55 1.08 0.78 1.55 PACAP + LPS 3.05 6.38 1.92 3.49 -
a BALB/c macrophages were preincubated for 2 h in the presence or absence of 10 ng/ml of PACAP 38. LPS (500 ng/ml) was added, and supernatants were collected after 4 h for TNF-α and IL-6 and tested by ELISA. Using the Mann-Whitney test, the decrease in TNF-α and increase in IL-6 when PACAP is preincubated with macrophages before adding LPS is significant, p < 0.05.
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